Pediatric cardiovascular disorders, which comprise congenital heart defects (CHD) and myocardial and conduction system diseases, remain highly challenging due to cardiac co-morbidities and premature mortality. As most of these disorders are genetic, efforts over the past 30 years have focused on identifying their causal mutations. Particularly for Mendelian traits such as Noonan syndrome and related disorders (the RASopathies), this has been highly successful. Newer genomic technologies have accelerated gene discovery for pediatric cardiovascular disorders, including genetically complex ones. These genetic discoveries are improving care through more accurate diagnosis, better prognostication, and refinement of clinical trial design. What has not occurred with rare exception is the development of novel therapies based on the new understanding of disease pathogenesis enabled by these gene discoveries. Finding therapies for pediatric cardiovascular disorders will be challenging because the biological targets are generally central to cell homeostasis (e.g., RAS/MAP kinase signaling) so cannot be completely inhibited for long periods without incurring side effects that would outweigh their benefits. For this R35 mechanism, I and my outstanding co- investigators with relevant expertise intend to address this gap using a drug development pipeline that begins with high-throughput screening to overcome pupal lethality in Drosophila melanogaster models of disease with a chemical library that covers druggable space (n=14,400) using 96-well plates and robotics. Screening in whole animals is performed agnostically and has the putative advantage of providing a simultaneous read out of efficacy and toxicity. We provide preliminary data showing that we have already achieved this using a fly RAF1 mutant model of Noonan syndrome with hypertrophic cardiomyopathy. Subsequent steps with fruit flies include confirmation of initial hits in vials, determining efficacy against adult fly phenotypes such as rough eye, ectopic wing veins and heart hypertrophy. Back-up libraries for the candidate compounds, typically 60-80 chemical neighbors, will be culled for ones with most desirable drug traits and then screened in the fly models. Using a defined set of fruit fly deficiency lines, targets and anti-targets will be established to enable further rounds of rational pharmacology. ADME studies will be used to reduce potential for drug-drug interactions. In parallel, we will pursue repurposing of FDA-approved drugs using library screening with fruit fly models and systems pharmacogenomics. Leading compounds and drugs will then be tested against phenotypes in human induced pluripotent stem cell lines with the disease-causing mutation for efficacy. The most promising drugs will then be tested in existing mouse models (e.g., HCM in Raf1 mutant mice) using appropriate endpoints. Taken as a whole, the approach proposed will significantly advance the identification of novel therapeutics for pediatric cardiovascular diseases, starting with the RASopathies and later for other traits. If robust, this will provide a paradigm that can be adopted for other genetic traits of interest to the NHLBI.